Treatments for diabetes mellitus and obesity

ABSTRACT

Methods of treatment in subjects suffering from diabetes mellitus or obesity are provided. The methods comprise the step of administering an active agent directly to the small intestine in the subject. In particular, the active agent may be administered directly to the duodenum in the subject. The active agents useful in the treatments described herein include analgesic agents and, in particular, antinociceptive agents such as capsaicin, resiniferatoxin, and their analogs.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a national phase application of PCT InternationalApplication No. PCT/US2012/047134, filed Jul. 18, 2012, which claims thebenefit of U.S. Provisional Application No. 61/509,491, filed on Jul.19, 2011, and U.S. Provisional Application No. 61/637,444, filed on Apr.24, 2012, the disclosures each of which are incorporated herein byreference in their entireties.

BACKGROUND OF THE INVENTION

Diabetes mellitus, or simply diabetes, is a metabolic condition, orcombination of conditions, in which an individual displays highconcentrations of blood glucose. The condition is caused either byinsufficient production of insulin within the body or by the failure ofcells to respond properly to the insulin that is produced.

Diabetes is one of the leading causes of death and disability in theUnited States and in other developed countries. Diabetes is associatedwith long-term complications affecting almost every part of the body.For example, the disease can lead to blindness, heart and blood vesseldisease, stroke, kidney failure, amputations, and nerve damage. Diabetesaffects approximately 8 percent of the United States population, and thetotal cost of diabetes in the United States alone is estimated toapproach $200 billion.

Diabetes is most typically classified as either type 1 diabetes (alsoreferred to as insulin-dependent diabetes or juvenile diabetes), whereinthe subject fails to produce sufficient insulin, type 2 diabetes (alsoreferred to as non-insulin-dependent diabetes, adult-onset diabetes, orobesity-related diabetes), wherein the subject fails to respond properlyto insulin, or gestational diabetes, wherein the condition develops inwomen late in pregnancy.

Type 1 diabetes results from the loss of insulin-producing beta cells inthe pancreas, generally due to an autoimmune response against the betacells. The cause of the autoimmune response is not known, but it isbelieved that genetic and environmental factors, including the possibleinvolvement of viruses, could be involved. Symptoms of type 1 diabetesinclude increased thirst and urination, hunger, weight loss, blurredvision, and extreme fatigue. Although it can appear at any age, type 1diabetes most frequently develops in children and young adults. Roughly5 to 10 percent of all diabetes cases are considered type 1 diabetes.

Type 2 diabetes is the most common form of diabetes, accounting for 90to 95 percent of all cases of diabetes. Type 2 diabetes is generallyassociated with older age, obesity, family history, previous historywith gestational diabetes, and physical inactivity. It is also moreprevalent in certain ethnicities. Type 2 diabetes is also referred to asinsulin-resistant diabetes because the pancreas is usually able toproduce sufficient amounts of insulin, but the body fails to respondproperly to that insulin. As with type 1 diabetes, blood glucose levelsin individuals suffering from type 2 diabetes increase, and the body isunable to metabolize the blood glucose efficiently. The symptoms of type2 diabetes generally develop more slowly than those of type 1 diabetes.The symptoms include fatigue, frequent urination, increased thirst andhunger, weight loss, blurred vision, and slow healing of wounds orsores. In some cases, no symptoms are evident.

Gestational diabetes occurs in approximately 3 to 8 percent of pregnantwomen in the United States, generally developing late in the pregnancy.The disease typically disappears after birth of the baby, but women whohave experienced gestational diabetes are significantly more likely todevelop type 2 diabetes within 5 to 10 years than those who have not.Women who maintain reasonable body weight and are physically activeafter suffering from gestational diabetes may be less likely to developtype 2 diabetes than those who do not. As with type 2 diabetes,gestational diabetes occurs more frequently among women with a familyhistory of diabetes and also in certain ethnic groups.

Since the discovery of insulin over 80 years ago, diabetes, particularlytype 1, or insulin-dependent diabetes, has been a somewhat treatablecondition. The combination of a proper diet, physical activity, andinsulin injection, together with the monitoring of blood glucose levelsusing portable meters, allows the management of type 1 diabetes. Fortype 2 diabetics, healthy eating, physical activity, and monitoringblood glucose levels are also important. In some cases, drug therapiescan be used to control blood glucose levels in these patients.

Other approaches to the treatment of diabetes are also desirable. Forexample, it has been shown that bariatric surgery for the treatment ofgross obesity may also cure obesity-related diabetes. It is thought thatthis surgery, which involves bypass of at least a portion of the smallintestine, could cause the surgical ablation of gastric inhibitorypolypeptide (GIP)-secreting intestinal K cells. Flatt (2007) Diab. Vasc.Dis. Res. 4, 150. As a result, circulating levels of GIP, which isthought to play a key role in lipid metabolism and fat deposition, aredecreased.

Furthermore, although the underlying cause of type 1 diabetes remainsunknown, a defect in a subset of sensory neurons that innervate thepancreas may play a role in initiating a series of events leading tolocal inflammation, destruction of islet cells in the pancreas, andautoimmune (type 1) diabetes. Razavi et al. (2006) Cell 127, 1123. Inindividuals with the defect, insulin secretion by beta cells fails toproperly stimulate sensory neurons expressing a hypofunctional,polymorphic capsaicin receptor (TRPV-1=transient receptor potentialvanilloid-1) to release neuropeptides. Suboptimal local levels ofneuropeptides lead to insulin resistance and beta cell stress andultimately an autoimmune response against the beta cells. Identifyingmethods to control these pathways could thus provide novel approachesfor treating or preventing diabetes.

Another health problem of major importance, particularly in thedeveloped world, is obesity. Obesity is a complex, multifactorial andchronic condition characterized by excess body fat. Obesity results froman imbalance between energy expenditure and caloric intake. Although thecauses of this imbalance are not completely understood, genetic and/oracquired physiologic events and environmental factors are important. Theadverse health effects of obesity, and more particularly morbid obesity,have become well-known in recent years. Such adverse health effectsinclude, but are not limited to, cardiovascular disease, diabetes, highblood pressure, arthritis, and sleep apnea. Generally, as a patient'sbody mass index (BMI) rises, the likelihood of suffering the adversehealth effects of obesity also rises.

Often, surgery has been the only therapy that ensures real results inpatients who have BMI values close to, or in excess of, 40 kg/m². Modernsurgical procedures generally entail either (1) the reduction of gastriccompliance, with the aim of limiting the subject's ability to ingestfood, or (2) the reduction of the food absorption surface by shorteningor bypassing part of the digestive canal. In some cases, both aims aresought through the same surgical procedure. The risk and invasivenessfactors of currently available surgeries are often too great for apatient to accept to undergo surgical treatment for treatment ofobesity. Accordingly, there is a need for less invasive, yet effectivetreatment procedures for the morbidly obese. Also, since the currentsurgical procedures are currently indicated only for those patientshaving a BMI of 40 or greater, or 35 or greater, when co-morbidities arepresent, it would be desirable to provide a less invasive procedure thatwould be available for slightly less obese patients, e.g., patientshaving a BMI of 30 to 35 who are not indicated for the currentlyavailable surgical procedures.

U.S. Patent Application Publication No. 2008/0275445 reports devices foruse in ablating tissue in the wall of luminal organs of thegastrointestinal tract. See also U.S. Pat. No. 7,326,207. Such devicesare purportedly useful in the treatment of pathophysiological metabolicconditions, such as type 2 diabetes, insulin resistance, obesity, andmetabolic syndrome, but the effectiveness of such treatments has notbeen demonstrated.

U.S. Patent Application Publication No. 2004/0089313 reports systems andmethods for treating obesity and other gastrointestinal conditions bytargeting the stomach. In particular, the methods purportedly causetightening of the pyloric sphincter or alteration of the receptiverelaxation of stomach muscles. The treatments include ablation ofstomach tissue and, in some cases, injection of a cytokine or avanilloid-containing compound into stomach tissue. The effectiveness ofthese treatments has not, however, been reported.

Given the widespread and growing significance of diabetes and obesitythroughout the population, there is clearly a strong need for additionalmethods of treating and preventing these diseases.

SUMMARY OF THE INVENTION

The instant disclosure addresses these and other problems by providingin one aspect methods of treatment comprising ablation of a region ofthe small intestine in a subject, wherein the subject suffers fromdiabetes mellitus or obesity.

In preferred embodiments, the region of the small intestine is a regionof the duodenum.

In some embodiments, the ablation targets a sensory nerve.

In some embodiments of the invention, the ablation is a mechanical,electrical, thermal, radiative, or chemical ablation. In specificembodiments, the ablation is a laser, ultrasound, radiofrequency,alternating current, microwave, or thermal ablation. In preferredembodiments, the ablation is a radiofrequency or alternating currentablation. In highly preferred embodiments, the ablation is a pulsedradiofrequency ablation.

In other highly preferred embodiments of the invention, the ablation isa chemical ablation. According to some of these embodiments, thechemical ablation may comprise administering an active agent directly tothe small intestine in a subject, wherein the subject suffers fromdiabetes mellitus or obesity.

In preferred embodiments, the active agent is administered directly tothe duodenum.

In some embodiments, the active agent causes ablation of sensory nerves.

According to some specific embodiments of the invention, the subjectsuffers from diabetes mellitus. In more specific embodiments, thesubject suffers from type 2 diabetes mellitus.

According to other specific embodiments of the invention, the subjectsuffers from obesity.

In preferred embodiments of the invention, the active agent administeredin the chemical ablation is an analgesic agent. In specific embodiments,the analgesic agent is an antinociceptive agent. In more specificembodiments, the antinociceptive agent is selected from the groupconsisting of capsaicin, a capsaicin analogue, resiniferatoxin, and aresiniferatoxin analogue.

According to some specific embodiments, the antinociceptive agent of thedisclosed methods is capsaicin or a capsaicin analogue. In otherspecific embodiments, the antinociceptive agent is resiniferatoxin or aresiniferatoxin analogue. In specific preferred embodiments, theantinociceptive agent is resiniferatoxin.

In some embodiments, the active agent used according to the methods ofthe invention is a ligand of the transient receptor potential cationchannel subfamily V, member 1, receptor.

In preferred embodiments, the active agent is administered by injection.In other preferred embodiments, the active agent is administered bytopical application.

According to some embodiments of the invention, the active agent isadministered by infusion through a syringe.

In specific embodiments, the active agent of the disclosed methods isadministered following surgical exposure of the small intestine of thesubject. In other specific embodiments, the active agent is administeredby a laparoscope or an endoscope.

In some specific preferred embodiments, the active agent of thedisclosed methods is administered by topical application directly to theduodenum, wherein the active agent is resiniferatoxin or aresiniferatoxin analogue, and wherein the patient suffers from diabetesmellitus. In other specific preferred embodiments, the active agent isadministered by topical application directly to the duodenum, whereinthe active agent is resiniferatoxin or a resiniferatoxin analogue, andwherein the patient suffers from obesity.

In some preferred embodiments, the methods of the invention furthercomprise measuring oral glucose tolerance of the subject.

In other preferred embodiments, the methods further comprise measuringbody weight change in the subject.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1. Effect of RTX treatment on glucose tolerance in rats. (A)Treatment on the surface of the duodenum; (B) Treatment on the surfaceof the ileum/colon; and (C) Infusion into the duodenum.

FIG. 2. Effect of RTX treatment on body weight in rats. (A) Treatment onthe surface of the duodenum; (B) Treatment on the surface of theileum/colon; (C) Infusion into the duodenum; and (D) Direct comparisonof the three RTX treatments.

DETAILED DESCRIPTION OF THE INVENTION

The present disclosure provides novel treatment methods for subjectssuffering from diabetes mellitus or obesity. Specifically, the methodsdisclosed herein comprise ablation of a region of the small intestine,preferably of the duodenum, and preferably wherein the ablation targetsa sensory nerve, in the subject.

As used herein, the duodenum is a specific segment of thegastrointestinal tract (GI tract). In certain instances the term GItract may refer to the “gut” or the “alimentary canal” that is acontinuous, coiled, hollow, muscular tube that winds through the ventralbody cavity. It is open to the external environment at both ends. In ahuman, its organs (gastrointestinal organs) generally include the mouth,pharynx, esophagus, stomach, small intestine (duodenum, jejunum, andileum), and large intestine (cecum, appendix, colon, rectum, and analcanal). The large intestine leads to the terminal opening, or anus.

The small intestine is the longest section of the digestive tract and isdivided into three segments: the duodenum, jejunum and ileum, each ofwhich performs different digestive functions. When the stomach contentsmove to the duodenum, the first segment of the small intestine, bile andpancreatic juice speed up digestion. Most of the iron and calcium in thefood we eat is absorbed in the duodenum. The jejunum and ileum, theremaining two segments of the nearly 20 feet of small intestine,complete the absorption of almost all calories and nutrients. The foodparticles that cannot be digested in the small intestine are stored inthe large intestine until eliminated.

Bariatric surgery produces weight loss by restricting food intake and,in some cases, by interfering with nutrition through malabsorption. Inparticular, bariatric surgeries in which all or part of the duodenum isbypassed are believed to cause a decrease in the absorption of food andthus result in weight loss in the patient. Furthermore, patientsundergoing gastric bypass surgery in which the duodenum is bypassed arefrequently cured of their type 2 diabetes. Pories et al. 1995, Ann.Surg. 222, 339. The exact physiological bases for these effects remainunknown, although studies in a non-obese diabetic rat model demonstratethat bypass surgery improves glycemic control with a significantdecrease in leptin and glucagon levels after oral glucose overload.Pacheco et al. 2007 Am. J. Surg. 194, 221. The effects of bypass surgeryon diabetes do not, therefore, result simply from surgically inducedweight loss or decreased caloric intake. More recent randomized clinicaltrials have demonstrated that gastric bypass surgery in obese patients,either in combination with intensive medical therapy or not, results inthe remission of type 2 diabetes in a significant number of thosepatients. Schauer et al. 2012 Mar. 26 N. Engl. J. Med. (epublication);Mingrone et al. 2012 Mar. 26 N. Engl. J. Med. (epublication). Bariatricsurgery is an extreme procedure, however, that is not typicallyindicated for non-obese diabetic patients. Alternative less costly andless invasive approaches to the treatment of diabetic and obese patientsare therefore desirable.

As described herein, the methods of the instant disclosure compriseablation of a region of the small intestine in a subject suffering fromdiabetes mellitus or obesity. The methods of the instant disclosure aresignificantly less invasive than, for example, bariatric surgery, andwill therefore result in lower morbidity and mortality in the treatedsubjects. The instant methods are also faster and less complicated thanalternative methods of treatment and will therefore result in costsavings compared to those methods.

In preferred embodiments of the instant invention, the ablation targetsa sensory nerve of the small intestine, or more particularly, of theduodenum. Without intending to be bound by theory, it is believed thatsuch sensory nerves may be involved in the loss of glycemic control indiabetic and obese patients and may therefore serve as desirable targetsfor ablation according to the methods of the instant invention. Ofparticular interest in the practice of the methods of the instantinvention is the targeting of sensory nerves that express the transientreceptor potential cation channel subfamily V, member 1 (TRPV-1)receptor. Such targeting can be accomplished, for example, byimmunolabeling or other labeling techniques during the ablationprocedure, as would be understood by those of ordinary skill in the art.In some embodiments of the invention, however, sensory nerves of thesmall intestine or duodenum are targeted more generally in the abalationstep, or they may not be targeted at all. In preferred embodiments ofthe invention, the targeted sensory nerve is on the serosal surface ofthe small intestine, most preferably on the serosal surface of theduodenum

Ablation of a region of the small intestine may be achieved according tothe instant methods using, for example, mechanical, electrical, thermal,radiative, or chemical ablation. In particular, laser, ultrasound,radiofrequency, alternating current, microwave, or thermal ablation maybe usefully employed in the practice of the instant methods.

In preferred embodiments, radiofrequency (RF) fields, such as, forexample, pulsed radiofrequency fields, may be usefully employed in theablation. Such pulsed radiofrequency fields are effective in reducingpain and other neuropathic syndromes, possibly by causingultrastructural changes in the neural structures to which they areapplied. The techniques appear to selectively target C-fibers andA-delta fibers, the principal sensory nociceptor axons, compared toA-beta fibers, which are related to touch and non-pain-relatedsensations. See, e.g., Erdine et al. 2009 Pain Pract. 9 407-417. PulsedRF techniques generate less heat in the target tissues than continuousRF techniques and may therefore operate by different mechanisms. Theamount of energy imparted by the RF device can be controlled anddirected, as would be known and understood in the art, in order toachieve the desired effects and to minimize side effects. Use ofselective ablation techniques, such as pulsed RF techniques, maximizesthe effectiveness of the treatments against diabetes and obesity whileminimizing side effects, such as, for example, those affecting tactilesensory inputs.

Radiofrequency ablation devices are well known in the surgical arts, andin particular in the art of cardiac surgery. See, e.g., U.S. Pat. Nos.4,945,912; 5,281,218; 5,348,554; and 5,423,808. Such devices can bereadily delivered to the desired location using a catheter or otherequivalent device. Radiofrequency ablation of the basivertebral nervehas recently been shown to reduce chronic back pain in clinical studiesusing devices designed to target intraosseous nerves. See, e.g., PCTInternational Publication No. WO 01/57655; U.S. Pat. No. 7,749,218.Analogous devices may be adapted for the practice of the methods of theinstant invention.

In other preferred embodiments of the invention, ablation of a region ofthe small intestine may be achieved using other electrical techniques,such as, for example, high-frequency alternating current stimulation.Such treatments have been shown to reversibly block the conduction of anaction potential through nerve cells. Variation in the frequency andamplitude of the stimulation provides selective effects on differentnerve fibers and thus allows for more control over the outcome of thetreatments, including the minimization of side effects. Specifically,the stimulation by high-frequency alternating current is selective inblocking slow-conducting, unmyelinated C-fibers, such as those ofnociceptive neurons, while minimizing effects on fast-conductingmyelinated A-fibers. See, e.g., Joseph et al. 2011 IEEE Trans NeuralSyst Rehabil Eng. 19 550-557. Such selectivity is useful in the methodsof the instant invention in targeting sensory neurons of the smallintestine and duodenum and thus in improving glycemic control and weightloss.

In some embodiments of the invention, it may be preferable for theablation to be reversible, whereas in other embodiments, it may bepreferable for the ablation to be irreversible. Adjustment andoptimization of the ablation technique to achieve the desired outcome,such as, for example, the recovery of glycemic control in a diabeticpatient or the loss of weight in an obese patient, are within the skillof the ordinary artisan.

In some embodiments of the invention, the ablation of a region of thesmall intestine is achieved using a chemical ablation technique. Inspecific embodiments, the methods comprise administering an active agentdirectly to the small intestine in a subject. The active agent usefullyadministered according to these methods is selected for itseffectiveness in treating diabetes mellitus and/or obesity upon directadministration. In particular, the active agent used in some of thedisclosed methods may cause ablation of sensory nerves in the smallintestine, and more specifically in the duodenum, and thus be effectivein treating these conditions.

In some embodiments, the active agent used according to some of theinstant methods may be an analgesic agent. Analgesic agents are widelyused and well known in the medical arts. Such analgesic agents may, forexample, be topical or systemic. Examples of topical analgesics arelidocaine and capsaicin. In some embodiments, the analgesic agent may bea psychotropic agent that has analgesic properties.

In preferred embodiments of the instant methods, the analgesic agent isan antinociceptive agent, specifically an agent that targets nociceptorson the surface of the small intestine, and in particular on the surfaceof the duodenum. In some embodiments of the instant methods, theantinociceptive agent is selected from the group consisting ofcapsaicin, a capsaicin analogue, resiniferatoxin, and a resiniferatoxinanalogue.

Resiniferatoxin and capsaicin and their analogues are exogenous ligandsfor the transient receptor potential cation channel subfamily V, member1 (TRPV-1), formerly known as the Vanilloid receptor 1 (VR1). The onlyknown endogenous ligand for this receptor is the proton. TRPV-1receptors are found on the visceral sensory nerve fibers of theunmyelinated type (C-fiber) and the thinly myelinated type (A-fiber).TRPV-1 functions as a modulator of sensory nerve responses to variousnoxious stimuli.

In some embodiments of the instant methods, the antinociceptive agent iscapsaicin or a capsaicin analogue. Capsaicin is the main pungentcomponent in hot pepper. Capsaicin treatment in animals can cause avariety of physiological effects. For example, systemic capsaicintreatment has been reported to alter serum glucose and free fatty acidconcentrations in rats and also to modulate the effects of insulin onthe levels of serum glucose and free fatty acids. At high levels,systemic capsaicin can cause the lesioning of afferent nerves inneonatal rats. Spiridonov et al. (2000) Neurosci. Behav. Physiol. 30,207. Such desensitization of sensory nerves can improve glucosetolerance and insulin sensitivity in treated animals. Desensitization bycapsaicin is also reported to delay the progression of type 2 diabetesin the Zucker fatty rat model. Gram et al. (2000) Diabetes 49, Suppl. 1,A428. It has further been reported that plasma levels of calcitoningene-related peptide (CGRP) are elevated in these rats prior to theonset of obesity and that the increased activity of sensory nervestherefore precedes the development of obesity and insulin resistance inthis model. Gram et al. (2005) Eur. J. Endocrinol. 153, 963.

Treatment of Zucker diabetic rats with systemic capsaicin reportedlytargets the expression of TRPV-1 receptors in CGRP-expressing primarysensory fibers in pancreatic islets of Langerhans cells and prevents thedeterioration of metabolism and glucose homeostasis in these animals.Gram et al. (2007) Eur. J. Neurosci. 25, 213. Furthermore, mutations inconserved regions of the TRPV-1 receptor have been identified innon-obese diabetic mice and have been correlated with neurogenicinflammation in islet beta cells. Razavi et al. (2006) Cell 127, 1123.Without intending to be bound by theory, TRPV-1 may therefore play arole in the development of both type 1 and type 2 diabetes through acombination of neuronal and autoimmune events. See also Bour-Jordan andBluestone (2006) Cell 127, 1097; Tsui et al. (2007) Trends Mol.Medicine. 13, 405.

TRPV-1 is highly expressed in sensory neurons, but it has also beendetected in mouse embryo 3T3-L1 preadipocytes and in human and mousevisceral adipose tissue. Zhang et al. (2007) Circ. Res. 100, 1063.Capsaicin is reported to induce calcium influx through the TRPV-1receptor in a dose-dependent manner in the preadipocyte cells and toprevent adipogenesis in stimulated cells. Adipose tissue from obesehumans reportedly displays decreased expression of the TRPV-1 receptor,and these cells likewise show a reduced capsaicin-induced calciuminflux. Studies in TRPV-1 knockout mice support a role for the receptorin adipogensis and obesity in vivo. Oral administration of capsaicin inmice was reported to prevent obesity in wild-type animals but not inTRPV-1 knockout mice fed a high-fat diet. Activators of TRPV-1 maytherefore prevent adipogenesis and obesity.

In some preferred embodiments, the antinociceptive agent of the instantmethods is resiniferatoxin or a resiniferatoxin analogue.Resiniferatoxin (RTX) is a naturally occurring super-analogue ofcapsaicin. It is found naturally in the latex of Euphorbia plants.Resiniferatoxin induces the desensitization effect and neuronal ablationmore potently than capsaicin and with minimal initial activation of theTRPV-1 neuron. Resiniferatoxin, and a number of agonists based on thestructures of capsaicin and resiniferatoxin have been reported aspotential analgesics through desensitization/denervation of theseneurons. RTX is reportedly less toxic than capsaicin when administeredsystemically.

Resiniferatoxin may be used to alleviate acute or chronic nausea,vomiting, or a combination thereof, by localized ablation of vagalafferent nerves in the stomach or other regions of the gastrointestinaltract. See U.S. Patent Application Publication No. 2009/0012154 A1, andreferences cited therein. It may also be used to ablate both painresponses and neurogenic inflammation. See U.S. Patent ApplicationPublication No. 2010/0204313 A1, and references cited therein.Furthermore, subcutaneous injection of resiniferatoxin in early diabeticrats may prevent further deterioration of glucose homeostatis, andadministration to overtly diabetic rats may improve glucose tolerance inthose animals. Gram et al. (2005) Eur. J. Pharm. 509, 211. Suchtreatments are likewise reported to improve the sensitivity of theanimals to insulin. Moesgaard et al. (2005) Am. J. Physiol. Endocrinol.Metab. 288 E1137.

RTX has been used to selectively ablate nociceptive neurons in thetreatment of hyperalgesia and neurogenic inflammation. Tender et al.(2005) J. Neurosurg. 102, 522. The effects of RTX on pain andinflammation occur without significant effects on normal tactilesensation or motor function. The treatments target nociceptive neuronsexpressing the TRPV-1 receptor and, in particular, the so-called C-fiberand A-delta fiber neurons. Since the majority of nerve fibers in theduodenum are C-fiber and A-delta fiber neurons, one of the features ofsome embodiments of the instant invention is to selectively targetsensory nerves of the small intestine and duodenum using capsaicin, RTX,and other ligands of the TRPV-1 receptor.

The chemical synthesis of resiniferatoxin has been reported. Wender etal. (1997) J. Am. Chem. Soc. 119, 12976.

Accordingly, in some embodiments of the instant methods, the activeagent used in the methods is a ligand of the TRPV-1 receptor. Asdescribed above, examples of such ligands include capsaicin and RTX.

The instant disclosure therefore also provides in another aspect aligand of the TRPV-1 receptor for use in treating diabetes mellitus orobesity. In preferred embodiments, the disclosure provides capsaicin,RTX, or the like, for use in treating diabetes mellitus or obesity.

In some embodiments of the instant methods, a single active agent may beadministered directly to the small intestine, and more specifically, tothe duodenum, in a subject. In some embodiments, combinations of activeagents may be used in the methods of treatment disclosed herein.

As recited in some of the instant methods, an active agent may beadministered directly to the small intestine in a subject. Directadministration of the active agent is in contrast to systemicadministration, in which an active agent is, for example, injected intothe bloodstream of a subject, is injected into the subjectsubcutaneously or intramuscularly, or is administered in any other waythat does not significantly localize the effects of the active agent tothe small intestine or duodenum. Direct administration may, for example,involve exposing the small intestine or duodenum directly to a stream ofsolution containing the active agent. In some embodiments, the activeagent may be administered to the small intestine or duodenum by topicalapplication. In some embodiments, tissues or organs adjacent to thesmall intestine or duodenum may be protected from exposure to the activeagent, for example, by the placement of sponges or other physicalbarriers. In some embodiments, the active agent may be administered by asyringe. In some embodiments, the active agent may be administered by acotton applicator, or other similar device. In some embodiments, theactive agent may be administered by another corresponding method.

According to some preferred embodiments of the instant methods, anactive agent may be administered directly to the serosal surface of theduodenum. As would be understood by those of ordinary skill in the art,the duodenum is comprised of layers, including the serosa, themuscularis, the submucosa, and the mucosa.

The active agent or agents of the instantly disclosed methods may beprovided in any manner that is effective in the treatment of a subjectsuffering from diabetes mellitus or obesity. For example, the agent maybe dissolved in a solution for direct administration to the smallintestine, and, in preferred embodiments, to the duodenum. The solutionmay contain other agents to, for example, stabilize or improvesolubility of the active agent. The solution may be an aqueous solutionor another solution capable of delivering the active agent to thesubject in an effective form. In some embodiments, as would beunderstood by the skilled artisan, the solution may be a mixture ofsolvents and may include other agents, such as, for example, detergents,such that the active agent is effectively delivered to the subject. Inpreferred embodiments, the active agent is dissolved in aqueoussolution. In other preferred embodiments, the solution also contains analcohol. In still other preferred embodiments, the solution alsocontains a detergent. In highly preferred embodiments, the solutioncontains both an alcohol and a detergent.

The active agent or agents of the instantly disclosed methods areprovided at concentrations effective to treat a subject suffering fromdiabetes mellitus or obesity. For example, the active agent may beprovided in a solution containing at least 1 ng, 3 ng, 10 ng, 30 ng, 100ng, 300 ng, 1 μg, 3 μg, 10 μg, 30 μg, 100 μg, 300 μg, 1 mg, 3 mg, 10 mg,30 mg, 100 mg, or even more active agent per mL solution. The activeagent may be provided in a solution containing at most 100 mg, 30 mg, 10mg, 3 mg, 1 mg, 300 μg, 100 μg, 30 μg, 10 μg, 3 μg, 1 μg, 300 ng, 100ng, 30 ng, 10 ng, 3 ng, 1 ng, or even less active agent per mL solution.In preferred embodiments, the solution contains the active agent fromabout 100 ng/mL to about 10 mg/mL solution. In more preferredembodiments, the solution contains the active agent from about 10 μg/mLto about 1 mg/mL solution. In even more preferred embodiments, thesolution contains the active agent at about 200 μg/mL.

Administration of active agents according to some methods of the instantdisclosure may, in some embodiments, be accomplished by surgicaltechniques. For example, the active agent may in some embodiments beadministered following surgical exposure of the small intestine of thesubject. Surgical exposure of the small intestine may be accomplished byany appropriate means, as would be understood by those of skill in theart.

Administration of active agents according to some methods of the instantdisclosure may, in some embodiments, not necessarily require thesurgical exposure of the small intestine of the subject. For example,the active agent may be administered in some embodiments by endoscopicor laparoscopic surgical techniques. Endoscopic or laparoscopicadministration may be accomplished by either using the endoscope orlaparoscope for both visualization as well as administration of theagent or by endoscopic or laparoscopic visualization to guideadministration of the agent.

General endoscopic procedural techniques such as visualizing, dilating,cutting, and manipulating tissue, and administering active agents, maybe accomplished using flexible devices such as endoscopes, balloons,snares and electrosurgical tools well known in the art. Laparoscopy orvisualization of the peritoneal cavity may be done, for example, with aflexible scope, as is typically used for diagnostic purposes, or with arigid scope, as is typically used for therapeutic procedures. With therigid scope procedures, a procedure is typically performed while beingobserved by secondary scopes positioned at various strategic positionsin the peritoneal cavity. In some embodiments, flexible scopes thatallow for visualization are not used for conducting the therapeuticprocedures by themselves, and the rigid scopes that have the toolsnecessary to perform the tasks associated with the therapeuticprocedures make use of additional flexible scopes so the doctor oroperator can view the tasks and procedure being performed. In someembodiments, such as traditional gastrointestinal endoscopy, tasks maybe performed with a scope that also allows the operator to view thetasks and procedures being performed. Gastrointestinal endoscopies mayin some embodiments be limited to work within the lumen or interiortissue of the gastrointestinal tract or associated organs, and not inthe peritoneal cavity. In other embodiments, the lumen of thegastrointestinal tract may, however, be breached by the endoscope inorder to reach surfaces exposed to the peritoneal cavity. See, forexample, U.S. Patent Application Publication No. 2007/0051380 A1. Insome embodiments, injection into the desired layers may be enhancedusing imaging guidance, such as by endoscopic ultrasound.

In some embodiments, the methods of the instant disclosure may furthercomprise the step of measuring oral glucose tolerance of the subject.Such measurement allows the efficacy of the treatment to be determined.Oral glucose tolerance may be measured by various methods, as would beunderstood by those skilled in the relevant art. For example, bloodglucose may be measured in a subject using a standard glucose meterfollowing the treatment methods. The blood glucose may be measured afterfasting to establish a baseline blood glucose level in the subject. Oralglucose may then be provided to the subject, and blood glucose atsubsequent times determined using the glucose meter to determine theoral glucose tolerance of the subject.

In some embodiments, the methods of the disclosure may further comprisethe step of measuring body weight change in the subject. Suchmeasurement may be helpful in determining the efficacy of treatment insubjects suffering from obesity and may, in some embodiments, be used incombination with the measurement of oral glucose tolerance in thesubject.

Further Aspects

In yet other aspects, the invention provides novel methods of treatmentaccording to the following numbered paragraphs:

1. A method of treatment comprising:

administering an active agent directly to the small intestine in asubject, wherein the subject suffers from diabetes mellitus or obesity.

2. The method of paragraph 1, wherein the active agent is administereddirectly to the duodenum.

3. The method of paragraph 1, wherein the active agent causes ablationof sensory nerves.

4. The method of paragraph 1, wherein the subject suffers from diabetesmellitus.

5. The method of paragraph 1, wherein the subject suffers from type 2diabetes mellitus.

6. The method of paragraph 1, wherein the subject suffers from obesity.

7. The method of paragraph 1, wherein the active agent is an analgesicagent.

8. The method of paragraph 7, wherein the analgesic agent is anantinociceptive agent.

9. The method of paragraph 8, wherein the antinociceptive agent isselected from the group consisting of capsaicin, a capsaicin analogue,resiniferatoxin, and a resiniferatoxin analogue.

10. The method of paragraph 8, wherein the antinociceptive agent iscapsaicin or a capsaicin analogue.

11. The method of paragraph 8, wherein the antinociceptive agent isresiniferatoxin or a resiniferatoxin analogue.

12. The method of paragraph 8, wherein the antinociceptive agent isresiniferatoxin.

13. The method of paragraph 1, wherein the active agent is a ligand ofthe transient receptor potential cation channel subfamily V, member 1,receptor.

14. The method of paragraph 1, wherein the active agent is administeredby injection.

15. The method of paragraph 1, wherein the active agent is administeredby topical application.

16. The method of paragraph 1, wherein the active agent is administeredby infusion through a syringe.

17. The method of paragraph 1, wherein the active agent is administeredfollowing surgical exposure of the small intestine of the subject.

18. The method of paragraph 1, wherein the active agent is administeredby a laparoscope or an endoscope.

19. The method of paragraph 1, wherein the active agent is administeredby topical application directly to the duodenum, wherein the activeagent is resiniferatoxin or a resiniferatoxin analogue, and wherein thepatient suffers from diabetes mellitus.

20. The method of paragraph 1, wherein the active agent is administeredby topical application directly to the duodenum, wherein the activeagent is resiniferatoxin or a resiniferatoxin analogue, and wherein thepatient suffers from obesity.

21. The method of any one of paragraphs 1-20, further comprising:

measuring oral glucose tolerance of the subject.

22. The method of any one of paragraphs 1-20, further comprising:

measuring body weight change in the subject.

It will be readily apparent to one of ordinary skill in the relevantarts that other suitable modifications and adaptations to the methodsand applications described herein may be made without departing from thescope of the invention or any embodiment thereof. Having now describedthe present invention in detail, the same will be more clearlyunderstood by reference to the following Examples, which are includedherewith for purposes of illustration only and are not intended to belimiting of the invention.

EXAMPLES Example 1 Effect of RTX on Blood Glucose Tolerance and BodyWeight in Rats

Resiniferatoxin (RTX) is a naturally occurring super-analogue ofcapsaicin. It is found naturally in the latex of Euphorbia plants.Capsaicin (CAP) is the main pungent component in hot pepper.Resiniferatoxin and capsaicin and their analogues are exogenous ligandsfor the transient receptor potential cation channel subfamily V, member1 (TRPV-1), which was formerly known as the Vanilloid receptor 1 (VR1).The only known endogenous ligand for this receptor is the proton. TRPV-1receptors are found on the visceral sensory nerve fibers of theunmyelinated type (C-fiber) and the thinly myelinated type (A-fiber).TRPV-1 functions as a modulator of sensory nerve responses to variousnoxious stimuli. Prolonged exposure to capsaicin leads to thedesensitization of the TRPV1 receptors and under certain situations,complete ablation of the nerves. Resiniferatoxin induces thedesensitization effect and neuronal ablation more potently thancapsaicin and with minimal initial activation of the TRPV-1 neuron.Resiniferatoxin, and a number of agonists based on the structures ofcapsaicin and resiniferatoxin have been reported as potential analgesicsthrough desensitization/denervation of theses neurons.

Experiments with systemic capsaicin or RTX have been shown to result inimprovement of type 1 and type 2 diabetes in animal model systems. Seeabove. Without intending to be bound by theory, the effects of theseagents may be due to a decrease in the release of the neurotransmittercalcitonin gene-related peptide (CGRP), which is characteristicallyproduced by sensory nerves and released upon stimulation. CGRP caninduce insulin resistance and inhibit insulin secretion in vitro.However, systemic exposure to RTX or capsaicin desensitizes sensorynerves everywhere and therefore is not desirable. The prior art isdeficient in failing to localize the effects of sensory nervedesensitization to a particular organ or to a particular surface of aparticular organ. As demonstrated herein, administering an active agent,such as RTX, capsaicin, or their analogues, directly to the smallintestine, in particular to the duodenum, in a subject improves glucosetolerance and causes weight loss. The results provided herein thusidentify for the first time a specific target for sensory nervedesensitization. The methods provided herein therefore allow for thesafe treatment of diabetes and obesity by directing, for example,capsaicin or RTX to a very restricted length of the intestine, therebyreducing or eliminating systemic side effects of these agents. Further,the methods provided herein can also be used to develop new methods forthe treatment of diabetes and obesity.

Methods

Animals

Adult male Sprague-Dawley rats (Harlan, Indianapolis, Ind.) weighing280-350 g were used in all the experiments. The animal protocol wasapproved by the Institutional Animal Care and Use Committee of theStanford University Medical Center.

Surgery and Resiniferatoxin (RTX) Treatment Rats were randomly dividedinto six groups: 1, vehicle solution treated on the surface of duodenum(n=4); 2, RTX solution treated on the surface of duodenum (n=4); 3,vehicle solution treated on the surface of ileum (1 rat) or colon (1rat) (n=2); 4, RTX solution treated on the surface of ileum (2 rats) orcolon (1 rat) (n=3); 5. Vehicle solution infuse into duodenum (n=2); 6.RTX solution infuse into duodenum (n=3).

Rats were fasted overnight prior to surgery having free access to water.Under 2% isoflurane-induced anesthesia, the abdomen was opened through amidline incision for the duodenum treatment and lower line for ileum orcolon treatment. These intestines were exposed and protected with warmsaline sponges around the intestine. Procedure 1: RTX application on thesurface of intestine: The cotton applicator with RTX (0.32 mM) orvehicle solution were gently applied topically to the outer surface ofentire duodenum (about 10 cm from the beginning of the duodenum) for 5mM Same treatment in ileum and colon groups were applied on about 10 cmlong of distal ileum and colon. Procedure 2: RTX or vehicle infused intoduodenum: Two atraumatic clamps used to block on beginning and distal ofduodenum. 0.8 ml of 0.32 mM RTX or vehicle was infused into duodenumthrough a glass syringe with a 27 gauge needle. After 30 mM the RTX orvehicle solution were withdrawn and the clamps were released. Aftertreatment, the abdominal incisions were closed.

RTX solution: RTX stock solution: 1 mg RTX+10% ethanol+10% Tween-80 insaline to make 1 μg RTX/μl in stock solution. RTX work solution (0.32mM): stock RTX solution (0.2 ml)+Saline (0.8 ml); Vehicle (like stocksolution without RTX)

Oral Glucose Tolerance Test (OGTT)

Post RTX treatment 1st, 3^(rd) and 7^(th) week all the rats wereperformed the OGTT. After 17 hours overnight fasting, blood glucose wasmeasured in conscious rats by tail-stick, using a glucose meter (BayerHealthcare LLC, Mishawaka, Ind.). OGTT was measured before (baseline)and each 15 minutes for 2 hours after oral gavage glucose (1 g/kg bodyweight).

Body Weight Change Monitoring

The body weight of rats was measured before RTX treatment and each bloodglucose testing day.

Data Analyses

Data are presented as means±s.e. Data were compared using Student'st-test. Differences were considered significant at <0.05.

Results

Oral Glucose Tolerance

RTX treated-rats on the surface of duodenum showed better glucosetolerance compared with vehicle group at 30 and 45-minute peak levels ofpost RTX treatment 1^(st), 3^(rd) and 7^(th) week (P<0.05 for both, FIG.1A). RTX treated-rats on the surface of ileum or colon and RTX infusinginto duodenum were not significantly different on glucose tolerance in 2hours testing time and all of testing day compared with vehicle groups(FIGS. 1B and C).

Body Weight Change

The body weight in rats treated on the surface of duodenum with RTX wassignificantly less in the 1^(st) week post RTX treatment compared withvehicle group (92.0±1.0 vs 98.8±2.0, P<0.05, FIG. 2A). After the 3^(rd)and 7^(th) week post RTX treatment, the body weight in both groups werenot significantly different. RTX treated-rats on the surface of ileum orcolon and RTX infusing into duodenum were not significantly different onbody weight of all measurement day compared with vehicle groups (FIGS.2B and C). On the other hand RTX treated-rats on the surface of duodenumshown also significantly loss of weight in 1^(st), 3^(rd) and 7^(th)week post RTX treatment compared with rats by RTX infusing into duodenum(92.0±1.0 vs 102.4±3.0, 110.5±2.6 vs. 123.1±1.6 and 118.9±0.9 vs.136.3±2.5, P<0.05, FIG. 2D).

All patents, patent publications, and other published referencesmentioned herein are hereby incorporated by reference in theirentireties as if each had been individually and specificallyincorporated by reference herein.

While specific examples have been provided, the above description isillustrative and not restrictive. Any one or more of the features of thepreviously described embodiments can be combined in any manner with oneor more features of any other embodiments in the present invention.Furthermore, many variations of the invention will become apparent tothose skilled in the art upon review of the specification. The scope ofthe invention should, therefore, be determined by reference to theappended claims, along with their full scope of equivalents.

What is claimed is:
 1. A method of treatment comprising: targetedchemical ablation of a sensory nerve of the duodenum in a subject,wherein the targeted chemical ablation comprises administeringresiniferatoxin, wherein the subject suffers from diabetes mellitus orobesity.
 2. The method of claim 1, wherein the chemical ablationcomprises administering the resiniferatoxin directly to a TRPV-1receptor.
 3. The method of claim 2, wherein the active agent isadministered by injection.
 4. The method of claim 1, wherein the activeagent is administered by topical application.
 5. The method of claim 2,wherein the active agent is administered by infusion through a syringe.6. The method of claim 1, wherein the active agent is administeredfollowing surgical exposure of the small intestine of the subject. 7.The method of claim 2, wherein the active agent is administered by alaparoscope or an endoscope.
 8. The method of claim 2, wherein theactive agent is administered by topical application directly to theduodenum, wherein the active agent is resiniferatoxin, and wherein thepatient suffers from obesity.
 9. The method of claim 1, wherein theagent resiniferatoxin is administered to the sensory nerve on theserosal surface of the duodenum.
 10. The method of claim 1, wherein thesubject suffers from diabetes mellitus.
 11. The method of claim 1,wherein the subject suffers from type 2 diabetes mellitus.
 12. Themethod of claim 1, wherein the subject suffers from obesity.
 13. Themethod of claims 1-12, further comprising: measuring oral glucosetolerance of the subject.
 14. The method of claims 1-12, furthercomprising: measuring body weight change in the subject.
 15. The methodof claim 1, wherein the sensory nerve expresses a transient receptorpotential cation channel subfamily V, member 1 (TRPV-1) receptor.